Cancer is a major cause of death worldwide, being the second-leading cause of death in developed countries and even the number one cause of death in e.g. Australia, Japan, Korea, Singapore and the male population of the UK and Spain. The number of people who develop cancer each year is increasing.
Increasing evidence points to the important roles of epigenetic alterations in cancer development. These involve DNA hypermethylation as well as chromatin modifications such as histone methylation and deacetylation (for a review see Feinberg, A. P., et al., Nature Reviews Genetics (2005) 7, 21-33). Many tumour suppressor genes have been found to be silenced by epigenetic mechanisms. Unlike genes harbouring disabling genetic mutations, tumour suppressor genes epigenetically silenced can be reactivated and to cause cells to go into apoptosis or senescence. This feature makes epigenetic modifications an ideal target for therapeutic interventions in cancer. It has been shown that specific inhibitors of DNA methylation, 5-azacytidine, and its deoxy analogue, 5-aza-2′-deoxycytidine can inhibit the DNA-dependent methyltransferase (DNMT) activity, reverse the silencing of tumour suppressor genes and have shown utility for the treatment of haematological malignancies. Clinical trials are also underway for agents that interfere with enzymes that modify histone, such as histone deacetylase inhibitors.
Similar epigenetic alterations such as DNA methylation and histone modifications appear to be involved in the maintenance, proliferation and differentiation of stem cells. Mature cells of the various tissues of an organism arise from progenitor cells, which possess a broad developmental potential and replicative capacity. Peripheral blood for example contains progenitor cells that can differentiate into endothelial or vascular smooth muscle cells. Progenitor cells in turn originate from stem cells, which have also been identified in most tissues. Stem cells are fully undifferentiated cells that are able to differentiate into any mature functional cells, such as heart, liver, brain cells etc., while retaining the ability to proliferate indefinitely. Methylation and demethylation of DNA are known to regulate transcriptional states during germ cell development, between fertilisation and blastocyst formation, and early development in mammals (for a review see e.g. Santos, F., and Dean, W., Reproduction (2004) 127, 643-651).
Evidence for a role of epigenetic alterations in the maintenance of stem cells is for instance the observation that late passages of embryonic stem cell cultures show a variety of clonal DNA alterations, such as a differential methylation of inter alia the promoter region of the putative tumour suppressor gene RASSF1 (Maitra et al., Nature Genetics (2005) 37, 10, 1099-1103). Epigenetic disruption of progenitor cells has furthermore been suggested to be a key determinant of cancer progression (Feinberg, A. P., et al., 2005, supra).
Currently, cancer therapy involves surgery or focuses on the functional or genetic changes associated with the transformation of cells into malignant cells. An ideal anti-cancer drug should selectively kill, or at least inhibit, rapidly proliferating cancerous cells, while leaving non-cancerous cells unaffected. Recent approaches include immunotherapy using antibodies directed to markers of selected types of cancer cells (e.g. US patent application 2005/0244417), the application of agonists to receptors that are expressed on certain types of cancer cells (US patent application 2006/0147456), the application of interferon-containing chitosan-lipid particles (US patent application 2005/0266093), as well as the application of a compound that acts as a cytotoxic agent for a certain type of prostate cancer cells by an unknown mechanism (US patent application 2005/0245559).
In recent years research efforts have also been undertaken to develop an epigenetic cancer therapy, since abnormal patterns of DNA methylation in cancer cells are known for more than 20 years (for an overview see e.g. Brown, R. and Strathdee, G., Trends in Molecular Medicine (2002) 8, 4 (Suppl.), S43-S48, or Yoo, C. B. and Jones, P. A., Nature Reviews Drug Discovery (2006) 5, 1, 37-50). Nevertheless only two DNA methyl-transferase inhibitors, 5-azacytidine (Vidaza™) and decitabine (Dacogen™) have made it to the market. They have been approved for the treatment of myelodysplastic syndrome, a haematological condition also known as “preleukemia”. There is therefore still a need in the art for novel compounds and compositions for treating or preventing cancer or neoplastic disease that preferentially kill rapidly cancerous cells.
With respect to the modulation of stem cells, the ability of embryonic stem cells to readily differentiate continues to pose a major practical challenge. In order to maintain embryonic stem cells in a pluripotent state, their differentiating during handling and growing in culture has to be prevented. For this reason they are traditionally cultured in the presence of fetal calf serum on a layer of feeder cells (see e.g. U.S. Pat. No. 5,843,780 and U.S. Pat. No. 6,090,622) or in fibroblast-conditioned medium (CM). Nevertheless, even under carefully controlled conditions embryonic stem cells may undergo spontaneous differentiation during in-vitro propagation. Leukaemia inhibitory factor, a factor mediating self-renewal in mouse embryonic stem cells, has also been found to inhibit differentiation of mouse embryonic stem cells, but it does not replace the role of feeder cells in preventing differentiation of human embryonic stem cells. Therefore, means of maintaining pluripotency and/or self-renewing characteristics of embryonic stem cells would be a substantial achievement towards realizing the full commercial potential of stem cell therapy.
Stem cells are also found in carcinomas called teratoma of various tissues (often of the testes and the ovary) that produce tissues consisting of a mixture of two or more embryological layers. The malignant forms of such carcinomas are also called teratocarcinoma. Development of stem cells in murine teratocarcinomas parallels events in the normal embryo. Their presence can explain that chemotherapy often removes the bulk of a tumour mass without preventing tumour recurrence (Chambers, I. Smith, A, Oncogene (2004), 23, 7150-7160). Furthermore, stem cells have been suggested to be a source of cancer cells in solid tumours, including breast and brain cancer. Leukaemias have been shown to be generated by a limited number of leukaemia stem cells of heterogeneous origin (Passegue, E. et al., Proc. Natl. Acad. Sci. USA (2003), 100, 11842-11849). Therefore, means of abrogating pluripotency and/or self-renewing characteristics of stem cells in such tumours would typically be a precondition for a permanent removal of such carcinomas.
Accordingly it is an object of the present invention to provide a method, as well as compounds and compositions that are capable of preferentially killing a cancer cell without affecting a non-cancerous cell. It is a further object of the present invention to provide a method of modulating pluripotency and/or self-renewing characteristics of a stem/progenitor cell.